Yersinia pestis, the causative agent of plague, is classified as a NIAID Category A Priority Pathogen due to its potential to be used as a biological weapon. The re-emergence of plague in countries where infection has remained dormant and the appearance of multidrug resistant strains indicate that Y. pestis remains a significant threat to human health. Y. pestis is primarily a pathogen of rodents, but humans can become infected through bite of an infected flea (causing bubonic plague) or through inhalation of respiratory secretions or aerosols containing Y. pestis (causing primary pneumonic plague). Despite the extensive history of humans and plague disease, little is known about Y. pestis pathogenesis at the molecular level. Autotransporter (AT) proteins represent a unique family of secreted virulence factors of Gram-negative bacterial pathogens, in that their primary sequences encode sufficient information to mediate secretion across both bacterial membranes. After secretion, ATs either remain tethered to the bacterial surface or are released via proteolytic cleavage. In silico analyses have identified ten classic ATs of Y. pestis (designated Yaps for Yersinia autotransporter proteins), two of which, YapE and YapC have been implicated in virulence. Furthermore, preliminary data suggest that another AT, YapG, is rapidly processed and released in Y. pestis and contributes to lymph node colonization during bubonic infection. Consequently, the long- term objective of this research proposal is to understand at a molecular level the role of YapG in bubonic and pneumonic disease. To this end, the goal of this research plan is to test the hypothesis that YapG plays an important role in Y. pestis pathogenesis by carrying out the following specific aims: (1) To assess the contribution of YapG to Y. pestis pathogenesis using the mouse models of bubonic and pneumonic infection (2) To determine the cleavage site(s) in YapG by mass spectrometry and examining the processing of mutant forms of YapG, and (3) To identify the potential function(s) of YapG by expressing YapG in Y. pestis and E. coli and determining whether it functions as a cytotoxin, adhesin or an autoagglutinin. Public Health Relevance: Although plague infection still remains a threat today with the existence of biological weapons and antibiotic resistant strains, no vaccine is available and little is known about the specific factors that contribute to disease. This research plan aims to study a protein, YapG, its contribution to Y. pestis pathogenesis, and how processing affects its potential function(s). By understanding how factors, such as YapG, contribute to plague disease, we can better design novel therapies and vaccine strategies to prevent Y. pestis infection.